Integrin activation during neutrophil adhesion and vascular inflammation

中性粒细胞对于宿主抵御细菌和真菌感染是必不可少的，它能诱导 组织坏死或缺血损伤后的炎症。如果不能正确解决，中性粒细胞 炎症是人类大量炎症性疾病背后的驱动力。中性粒细胞到达 先打滚，然后抓捕，从血液中受到伤害的地方。停滞是由趋化因子触发的 诱导了..2整合素的高亲和力构象。两个FERM结构域蛋白，kindlin-3和 Talin-1，是中性粒细胞抑制所必需的。众所周知，Talin-1是一种结合TALIN-1的接头蛋白 肌动蛋白细胞骨架，通过结合和改变整合素的拓扑结构来激活整合素。跨膜 域。我们最近报道了kindlin-3通过其pleckstrin被募集到质膜上。 中性粒细胞停止之前的同源结构域。然而，中性粒细胞扩散和 Kindlin-3在这一过程中的作用还知之甚少。此外，kindlin-3与talin-1是如何协同作用的 激活整合素，以及Kindlin-3是否也通过直接与肌动蛋白结合而发挥接头蛋白的功能 是完全未知的。我们的主要假设是，在中性粒细胞扩散的过程中，两者都激活了kindlin-3 和talin-1同时被招募到质膜，在那里kindlin-3组织高亲和力 整合素激活。为了验证这一假设，我们产生了用于同时检测的报告鼠系 整合素活化和小鼠中性粒细胞中kindlin-3和talin-1的成像。在目标1中，我们将测试 假设Kindlin-3在中性粒细胞扩散过程中组织了一环聚集的高亲和力整合素环 在流动条件下；在目标2中，我们将测试kindlin-3通过以下方式调节整合素激活的假设 直接与肌动蛋白细胞骨架结合；在目标3中，我们将确定..2整合素激活的作用。 在缺血再灌注损伤(IRI)中。拟议的研究在概念上是创新的，并且是高度的。 意义重大，因为它将解开kindlin-3如何组织高亲和力整合素激活的谜团 并确定它在IRI诱导的炎症中所起的作用。成功完成这项提案将 建立整合素激活的分子机制并提供对中性粒细胞的机制洞察 扩散和血管炎症。

Neutrophils, which are essential for host defense against bacterial and fungal infections, induce inflammation following tissue necrosis or ischemic injury. If not properly resolved, neutrophilic inflammation is the driving force behind a plethora of human inflammatory diseases. Neutrophils arrive at the site of injury from the bloodstream by first rolling and then arresting. Arrest is triggered by chemokines that induce the high-affinity conformation of ..2 integrins. Two FERM domain proteins, kindlin-3 and talin-1, are required for neutrophil arrest. It is well known that talin-1, an adaptor protein that binds the actin cytoskeleton, activates integrins by binding and altering the topology of the .. transmembrane domain. We recently reported that kindlin-3 is recruited to the plasma membrane through its pleckstrin homology domain prior to neutrophil arrest. However, the mechanism underlying neutrophil spreading and the role of kindlin-3 in this process are poorly understood. Moreover, how kindlin-3 cooperates with talin-1 to activate integrins and whether kindlin-3 also functions as an adaptor protein by directly binding to actin are completely unknown. Our overarching hypothesis is that, during neutrophil spreading, both kindlin-3 and talin-1 are simultaneously recruited to the plasma membrane, where kindlin-3 organizes high-affinity integrin activation. To test this hypothesis, we generated reporter mouse lines for simultaneous detection of ..2 integrin activation and imaging of kindlin-3 and talin-1 in mouse neutrophils. In Aim 1, we will test the hypothesis that kindlin-3 organizes a ring of clustered high-affinity ..2 integrins during neutrophil spreading under flow conditions; in Aim 2, we will test the hypothesis that kindlin-3 regulates integrin activation by directly binding to the actin cytoskeleton; and in Aim 3, we will determine the role of ..2 integrin activation in ischemia-reperfusion injury (IRI). The proposed research is conceptually innovative and highly significant because it will resolve the enigma of how kindlin-3 organizes high-affinity integrin activation and define the role it plays in IRI-induced inflammation. Successful completion of this proposal will establish molecular mechanisms of integrin activation and provide mechanistic insight into neutrophil spreading and vascular inflammation.